High-Throughput Method for Lipophilicity Measurement

Abstract

A high-throughput method has been developed for lipophilicity measurement. It measures the partition coefficient of a solute between a polymer phase and an aqueous phase (P pw ) in a 96-well format. The polymer is plasticized poly(vinyl chloride) (PVC), which is widely used as a material for clinical containers and ion-selective electrodes. The composition is 2:1 (w/w) dioctyl sebacate (DOS) and PVC. With 6 repeats, log P pw values of 15 solutes have been determined in one 96-well microplate in 4 hours. A linear relationship between log P pw and log P ow (octanol-water partition coefficient) values exists with a correlation coefficient of 0.979. The slope and intercept of the log P pw vs. log P ow plot are statistically indistinguishable from 1 and 0, respectively. Similar to the HPLC method, by using the correlation line as a calibration curve, the measured log P pw values can be used to predict log P ow . This protocol is faster than the HPLC method. Moreover, it is straightforward to extend the protocol to the determination of the distribution coefficient and pK a of charged solutes. We show that the log P pw of the neutral form of racemic econazole is 4.83(±0.06), for the cationic form (presumably as a dihydrogen phosphate ion pair) 1.68(±0.04), and the pK a is 6.15(±0.04). This method has great flexibility as well, and is potentially fully automated.Lipophilicity represents the affinity of a molecule or a moiety for a lipophilic environment. 1, 2 Over a century ago, Overton 3 and Meyer's 4 pioneering work demonstrated that the lipophilicity of a compound could be related quantitatively to its biological activity. As suggested by Collander 5 and Hansch, 6, 7 the logarithmic value of the n-octanol/water partition coefficient, log P ow , has been widely recognized in the pharmaceutical, biomedical and environmental sciences to describe the lipophilicity of various compounds. Numerous experimental methods exist to measure log P ow values. The shake-flask procedure is a standard method 8 to determine log P ow in the range of -2 to 4, but it is time and labor consuming, and requires relatively large amounts of pure compounds. 9, 10 In addition, entrainment and octanol/water emulsions can be severe problems for compounds having a log P ow value larger than 4. 11 The HPLC method is an indirect way to estimate log P ow values in the range of 0 to 6, and has also become a standard procedure. 12, 13 A series of reference compounds is injected onto a C18 column. The compounds' retention factors are used to create a calibration curve with their known log P ow values. Compounds with unknown log P ow values are then injected and their retention factors are used to predict log P ow from the calibration curve. However, this technique is only valid for neutral molecules since charged molecules have a far more complex retention behavior than simple partitioning. HPLC and other separation-based methods to determine log P ow have been reviewed. 11, 14, 15 Responding to the need for more speed and accuracy, workers have ...

Determination of binding constants by affinity capillary electrophoresis, electrospray ionization mass spectrometry and phase-distribution methods

“…Recently, a high-throughput phase-distribution method was introduced for the determination of octanol/water partitioning [109], chirally discriminating binding [105] and drug-cyclodextrin binding [110]. Taking the last study as an example of the approach, as shown in Fig.…”

Lipophilicity screening of novel drug-like compounds and comparison to clogP

“…It is known that predictions are not particularly good for partition coefficients and pK a values of zwitterionic, tautomeric and charged compounds[8]. The UHPLC method gives better sensitivity than the optical absorbance measurements[10]. Unlike the plate reader method, it does not suffer from the potential for interfering compounds biasing the result.…”

“…Unlike the plate reader method, it does not suffer from the potential for interfering compounds biasing the result. The higher sensitivity translates into the method’s capability to measure a wider dynamic range of log P pw values in comparison to the optical absorbance approach [10]. On the other hand, compared to the plate reader, the UHPLC measurement generates lower throughput due to method development time.…”

“…The polymer phase is composed of poly (vinyl chloride) (PVC) and dioctyl sebacate (DOS) at the ratio of 1:2 (w/w). Studies in our group[10] on the correlation of polymer/water partition coefficient log P pw and standard 1-octanol/water partition coefficient (log P ow ) have shown a good linear relationship with the slope of 0.933 and the intercept close to zero, indicating that DOS plasticized PVC had lipophilicity similar to octanol. Therefore, our polymer/water partition coefficient can be used to predict lipophilicity.…”

Estimating the Lipophilicity of Natural Products Using a Polymeric Reversed Phase HPLC Method

“…[4, 5, 6] All of these methods achieve satisfactory correlations with Log P oct , even for very hydrophobic compounds (Log P oct > 6). Valkó et al proposed a rapid method for measurement of Log P via reversed-phase HPLC and established a general solvention equation for Log P values and Chromatographic Hydrophobicity Indices with acetonitrile (CHI ACN ).…”